PROJECT SUMMARY/ABSTRACT Generally, Prostate Cancer (PCA) is a slow-growing malignancy with decades of indolence, but takes on an aggressive form displaying rapid growth, dissemination, and lethality in a subset of cases (< 20%). Consequently, a critical unmet need exists in developing prognostic tools to identify aggressive PCA foci among non-aggressive foci. The clinical management of PCA is challenging since the current methods for active surveillance to distinguish aggressive lesion from non-aggressive lesion from unifocal or multifocal tumors. The goal is to be able to identify aggressive lesions from multifocal lesions in PCA patients. This will benefit patients with indolent PCA in avoiding radical prostatectomy as well its side effects. The long-term goal is to determine tumor aggressiveness at an early stage using DNA methylation biomarkers in order to guide clinical decisions on focal ablation therapy. The objective in this particular application is to identify DNA methylation markers that can improve the diagnostic value of prostate needle biopsy. Our central hypothesis is that combining specific DNA methylation marker analyses of needle biopsies with histological data will improve diagnostic accuracy for potentially aggressive unifocal or multifocal prostate cancer lesions and active surveillance. The rationale for the proposed research is that since changes in DNA methylation can occur prior to histological changes and can be detected in small amounts of tissue, DNA methylation markers in needle biopsy material would be able to identify not only prostate cancer, but also aggressive lesions in a multifocal environment. We will test this hypothesis by pursuing two specific aims: 1) to confirm whether DNA methylation alterations can be used to identify aggressive PCA lesions, either focally or multifocally, in primary clinical specimens. These markers will be further validated in the cohort from The Cancer Genome Atlas (TCGA) database; 2) to evaluate the identified DNA biomarker signatures in Aim 1 for their clinical applicability in the USC Prostate Biopsy Sample Repository using a cohort of ultrasound-guided needle biopsy samples from unifocal or multifocal tumors. We will then compare the DNA methylation marker results in needle biopsy to the histopathological profiles and patient outcome in order to evaluate the sensitivity and specificity of this combinatorial approach compared to using DNA methylation markers alone. The approach is innovative because it represents a significant departure from the current method of characterizing PCA aggressiveness. The proposed research is significant because it is expected to considerably increase the diagnostic accuracy of prostate needle biopsies. Ultimately, the use of DNA methylation markers in diagnosing aggressive PCA will improve patient quality of life and reduce the overall health burden by limiting surgical intervention while promoting focal treatment of prostate cancer.